Blondelle S E, Pérez-Payá E, Allicotti G, Forood B, Houghten R A
Torrey Pines Institute for Molecular Studies, San Diego, California 92121, USA.
Biophys J. 1995 Aug;69(2):604-11. doi: 10.1016/S0006-3495(95)79934-9.
A clear understanding of the specific secondary structure and binding domain resulting from the interactions of proteins and peptides with lipid surfaces will provide insight into the specific functions of biologically active molecules. We have shown in earlier studies that the stationary phases used in reverse-phase high-performance liquid chromatography represent a model artificial lipid surface for the study of induced conformational states of peptides on lipid interaction. We have now used reverse-phase high-performance liquid chromatography to determine the binding domains of peptides and, by extension, of proteins to a lipid surface. This approach consists of performing chemical modifications of specific amino acid side-chain functionalities after the interaction of the peptides with the reverse-phase high-performance liquid chromatography C18 groups. The susceptibility to oxidation was also studied after binding of the same peptides to liposomes. Oxidation of a single methionine residue "walked" through an amphipathic alpha-helical 18-mer peptide was selected to illustrate this approach. The extent of oxidation was found to be clearly dictated by the accessibility of the methionine residue to the aqueous mobile phase. The binding domain found for the peptide in its lipid-induced conformational state was unequivocally the entire hydrophobic face of the amphipathic alpha-helix.
清楚了解蛋白质和肽与脂质表面相互作用所产生的特定二级结构和结合域，将有助于深入了解生物活性分子的特定功能。我们在早期研究中表明，反相高效液相色谱中使用的固定相代表了一种模型人工脂质表面，用于研究肽在脂质相互作用时诱导的构象状态。我们现在已使用反相高效液相色谱来确定肽以及由此推广到蛋白质与脂质表面的结合域。该方法包括在肽与反相高效液相色谱C18基团相互作用后，对特定氨基酸侧链官能团进行化学修饰。在相同肽与脂质体结合后，还研究了其氧化敏感性。选择对一条两亲性α-螺旋18聚体肽中单个甲硫氨酸残基的氧化“游走”情况进行研究，以说明该方法。发现氧化程度明显取决于甲硫氨酸残基与水相流动相的可及性。在脂质诱导的构象状态下，该肽的结合域明确为两亲性α-螺旋的整个疏水表面。
